Early- and late-phase estrogenic responses in the uterus have been recognized for more than 60 years, yet mechanisms involved in their regulation remain controversial. One concept is that an early events(s), occurring within the first 6 h, prepares the uterus for later (18-30 h) increase in DNA synthesis, cell proliferation and protein synthesis. An alternate view is that the late growth phase is a result of the continuous presence of a stimulus. Discussion of either concept usually makes the assumption that all of the responses are dependent upon ligand interaction with one of the two estrogen receptor isoforms (ER1 and ER2). However, increased gene expression following injection of estrogen or xenoestrogen, in mice lacking ER1, or in which all ER-activity has been suppressed by an estrogen-antagonist, ICI 182,780 (ICI), has shown this to be an oversimplification. In this regard, accumulating evidence suggests that estrogen regulates diverse but interdependent signaling pathways in uterine biology via ER- dependent and -independent manners. While our long standing hypothesis is that estrogenic certain early responses are ER-independent, late responses are ER1- dependent, and a cross-talk between the two phases is necessary for a full complement of estrogenic responses in the uterus. Our previous and preliminary observations suggest that among several such early genes, Bip and Sik-SP are induced by estradiol- 172 (E2) and kepone in the mouse uterus via ER-independent manner as a phase-I response. Moreover, we recently showed that Bip is critically necessary in mediating E2- or kepone-dependent estrogen signaling that involves ER1 functions. Our preliminary studies also indicated that E2 induces GPR30 and ERK1/2 phoshorylation without involving ER1. Furthermore, we observed that the administration of a selective inhibitor of ERK1/2 activation (SL327), 30 min prior to E2 injection abrogates early ER- independent genes, as well as uterine wet weights during the early and late responsive phases. In contrast, injection of the inhibitor 6 h after E2 injection did not alter late effects. Collectively, these results positioned us to study early and late estrogenic responses to determine molecular relationship between the phases. Our specific aims are to elucidate: 1. Early molecular signaling by E2 or kepone involving Sik-SP, Bip and GPR30 in the uterus. 2. Functional significance of Sik-SP and GPR30 in E2- or kepone- mediated late estrogenic responses in the uterus. Studies should yield new molecular insights involving Bip, Sik-SP and GPR30 during ER1-mediated estrogen signaling. Overall, the results will help us to define relationship and a molecular cross-talk between the two-phase estrogenic responses in the uterus. PUBLIC HEALTH RELEVANCE: Our specific aims are to elucidate: 1. Early molecular signaling by E2 or kepone involving Sik-SP, Bip and GPR30 in the uterus. 2. Functional significance of Sik-SP and GPR30 in E2 or kepone mediated late estrogenic responses in the uterus. Studies should yield new molecular insights involving Bip, Sik-SP and GPR30 regulation of ER1-mediated estrogen signaling. Overall, the results will help us to define the relationship and a molecular cross-talk between the two-phase estrogenic responses in the uterus.